Kinase Domain Point Mutations in Ph+ Acute Lymphoblastic Leukemia (ALL) and Lymphoid Blast Crisis of Chronic Myeloid Leukemia (CML) and Their Emergence Following Therapy with Bcr-Abl Kinase Inhibitors.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1831-1831
Author(s):  
Dan Jones ◽  
Rayjalakshmi Luthra ◽  
Hagop M. Kantarjian ◽  
Megan Breeden ◽  
Susan O’Brien ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Direct Sequencing ◽  
Lymphoblastic Leukemia ◽  
Blast Crisis ◽  
Philadelphia Chromosome ◽  
Point Mutations ◽  
Kinase Domain ◽  
Current Therapy ◽  
Abl Kinase ◽  
T315i Mutation

Abstract Bcr-Abl kinase domain (KD) point mutations are detected in the dominant clone(s) in approximately 45% of CML at the time of disease resistance, developing after an average of 20–35 months of imatinib therapy. However, low numbers of Philadelphia chromosome (Ph)+ tumor cells with KD mutations could be present at earlier timepoints providing a pool of potential resistant subclones. Since current therapy of Ph+ ALL relies on imatinib maintenance therapy, the pattern of Bcr-Abl KD mutations in this tumor is an important and understudied phenomenon. We assessed the frequency and levels of Bcr-Abl KD mutations at different points in ALL, including at diagnosis, upon relapse and following salvage therapy with kinase inhibitors. We performed Bcr-Abl KD mutational analysis by direct sequencing in 25 cases of Ph+ ALL at the time of diagnosis and 25 cases upon disease persistence/relapse. For comparison, we analyzed 22 cases of lymphoid blast crisis of CML (LyBC), most of which transformed following long-term imatinib monotherapy. To track the emergence of mutated clones, we also performed more sensitive analysis for the T315I mutation by pyrosequencing (5% sensitivity) and allele-specific oligonucleotide probe (ASO) PCR (1:500 sensitivity). KD mutations were not seen by direct sequencing in ALL cases at diagnosis. The T315I mutation was also not detected by pyrosequencing (n =25) or ASO-PCR (n = 10) in newly diagnosed ALL. In contrast, Bcr-Abl KD mutations (Y253H in 3, Q252H, T315I, F317L, E355Q, H396R in 1 each) were seen in 8 of 25 (32%) relapsed/persistent ALL, occurring in patients who had been receiving imatinib for a median of 14 months (range 2–26). An additional 3 patients treated with dasatinib or nilotinib for relapse subsequently developed KD mutations (T315I and Y253H, and F317L) after 1, 4 and 9 months of second therapy. KD mutations were seen in 16 of 22 (73%) patients with lymphoid blast crisis, including T315I in 7, E255K and M244V in 2 each, and Y253H, V299L, F311I, E355G, F359V in 1 each. All KD mutations in LyBC developed following imatinib or nilotinib therapy. As with CML, kinase inhibitor therapy particularly in the relapse/salvage setting is the primary risk factor for emergence of Bcr-Abl KD mutations in Ph+ ALL. There is a high frequency of Bcr-Abl KD mutations associated the lymphoid transformation of CML. However, Bcr-Abl KD mutations develop more rapidly in persistent or relapsed Ph+ ALL than in CML and there is a higher frequency of Y253H mutations noted. These findings will likely have consequences for the timing and dosages of imatinib and other kinase inhibitors in maintenance and relapsed ALL regimens.

Combination Chemotherapy With Tyrosine Kinase Inhibitors Can Overcome Bcr-Abl Mutations In Acute Lymphoblastic Leukemia Or Blast Crisis CML

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2734-2734
Author(s):  
Ohad Benjamini ◽  
Hagop M Kantarjian ◽  
Susan O'Brien ◽  
Deborah Thomas ◽  
Farhad Ravandi ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Tyrosine Kinase ◽  
Tyrosine Kinase Inhibitors ◽  
Myeloid Leukemia ◽  
Kinase Inhibitors ◽  
Lymphoblastic Leukemia ◽  
Blast Crisis ◽  
Point Mutations ◽  
Kinase Domain ◽  
T315i Mutation

Background Point mutations in the kinase domain of bcr-abl confer resistance to tyrosine kinase inhibitors (TKIs) in patients with blast phase chronic myeloid leukemia (CML-BC) and Philadelphia positive acute lymphoblastic leukemia (Ph+ ALL). In particular the presence of T315I mutation is highly resistant to most current available TKIs.

Screening of Mutations in BCR-ABL Kinase Domain in Chronic Myeloid Leukemia (CML) Patients Treated with Kinase Inhibitors by Denaturing High-Performance Liquid Chromatography (D-HPLC).

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4580-4580
Author(s):  
Cintia C. Mascarenhas ◽  
Anderson F. Cunha ◽  
Katia B.B. Pagnano ◽  
Rosana A. Silveira ◽  
Fernando F. Costa ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Transplantation ◽  
Marrow Transplantation ◽  
Kinase Inhibitors ◽  
Blast Crisis ◽  
Point Mutations ◽  
Kinase Domain ◽  
Molecular Response ◽  
Abl Kinase ◽  
Clinical Resistance

Abstract Point mutations within the ABL kinase domain are the most frequent mechanism for reactivation of kinase activity of the BCR-ABL gene and have been associated with clinical resistance to tyrosine kinases (TK) inhibitors in CML patients conferring in some of them a poor prognosis. The T315I (Treonine → Isoleucine) is a mutation described in exon 6 of BCR-ABL gene that makes the protein resistant to all kinase inhibitors most currently used for treating CML (imatinib, nilotinib and dasatinib). D-HPLC allows for high throughput mutation screening. This technique is based on heteroduplex formation by PCR products amplified from wild type and mutant alleles. Under optimized denaturing conditions, these heteroduplexes can be distinguished from homoduplex. In this study we screened mutations in exon 6 of BCR-ABL gene in patients treated with kinase inhibitors, in different phases of the disease. We evaluated 85 patients: 9 at diagnosis, 81 in chronic phase, 3 in accelerated phase, one in blast crisis. Thirty four were resistant to imatinib, 10 of them to dasatinib and three had suboptimal response to imatinib. In 9 of 85 (10,5%) samples, D-HPLC showed an abnormal elution profile suggesting the presence of nucleotide changes. Automated sequencing confirmed the presence of two point mutations: T315I (two patients) and F359V (two patients). Five patients requires sequencing confirmation. Patients with T315I mutation failed to imatinib and dasatinib. One of them relapsed after bone marrow transplantation in blast crisis. Patients with F359V mutation were resistant to imatinib. One of them has partial hematological response with dasatinib and the other is in complete molecular response after bone marrow transplantation. D-HPLC seems to be a ship and practical method for routine clinical monitoring for emergence of kinase domain mutations and may be useful for optimizing therapy in CML. Early detection of emerging mutant clones may help in decision-making of alternative treatment.

Pathobiology of Lymphoid and Myeloid Blast Crisis and Management Issues

Hematology ◽  
2005 ◽  
Vol 2005 (1) ◽  
pp. 188-194 ◽  
Author(s):  
Robert L. Ilaria
Keyword(s):  
Kinase Inhibitors ◽  
Early Stage ◽  
Blast Crisis ◽  
Clonal Evolution ◽  
Philadelphia Chromosome ◽  
Kinase Domain ◽  
Early Stage Disease ◽  
Abl Kinase ◽  
Road Map ◽  
Blastic Phase

Abstract Despite recent improvements in the treatment of early-stage disease, the blastic phase of chronic myeloid leukemia (CML) remains a therapeutic challenge. For imatinib-naïve patients, imatinib provided encouraging hematologic and cytogenetic benefits; however, the vast majority of CML blast crisis cases today arise in patients already on imatinib-based therapy. Clonal evolution and duplication of the Philadelphia chromosome continue to be associated with blastic phase transformation, but recent studies have identified BCR/ABL kinase domain mutations in 30%–40% of blast crisis patients. This implies that BCR-ABL–targeted therapy might have influenced the molecular road map to blastic transformation. In this review, we will examine the effect of imatinib on primitive CML progenitors and how this might influence the pathophysiology of blast crisis. A rational framework for deciding how best to integrate stem cell transplantation, traditional chemotherapy, imatinib, and other BCR-ABL kinase inhibitors in the care of blast crisis patients will also be discussed.

Dynamics and Characteristics of BCR-ABL Multiple Mutations In Tyrosine Kinase Inhibitor Resistant Chronic Myeloid Leukemia.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3443-3443
Author(s):  
Dong-Wook Kim ◽  
Dongho Kim ◽  
Soo-Hyun Kim ◽  
Saengsuree Jootar ◽  
Hyun-Gyung Goh ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Point Mutation ◽  
Clinical Relevance ◽  
Kinase Inhibitors ◽  
Direct Sequencing ◽  
Point Mutations ◽  
Kinase Domain ◽  
Abl Kinase ◽  
Multiple Mutation ◽  
Compound Mutation

Abstract Abstract 3443 BCR-ABL kinase domain (KD) point mutation causes resistance to tyrosine kinase inhibitors (TKI) in CML patients through impaired binding of TKI to the target site. One of the characteristics of patients with BCR-ABL kinase domain point mutations is the fact that some patients have multiple mutations. However there have not been many studies showing that data about clinical relevance or dynamics of multiple mutation during CML treatment. From January 2002 to June 2010 at Seoul St Mary's Hospital, 277 CML patients were screened for mutation analysis due to sign of resistance to tyrosine kinase inhibitors including imatinib, nilotinib, dasatinib or bosutinib. We found that 95 patients have point mutation in BCR-ABL kinase domain through direct sequencing or ASO-PCR. Among them, 17 patients showed multiple mutation containing more than one type of point mutations in BCR-ABL KD. We investigated the patients with multiple mutations to characterize its clinical relevance and dynamics. Once mutation found, follow-up samples from the corresponding patients were collected and analyzed prospectively, or mutation status was analyzed retrospectively with cryopreserved samples if they were available. Status of the patients with multiple mutation is shown in Table 1. In order to investigate whether the multiple mutations are on same clone or on separated clone, we cloned serial samples from the 17 patients. Cloning of cDNA region corresponding to BCR-ABL KD into plasmid was performed and followed by transformation into competent cells, colony formation, plasmid preparation of 20 colonies from each sample, and then direct sequencing. Multiple mutations of 88% patients (15 out of 17) existed compound mutation which means the individual mutant types are located on the same BCR-ABL molecule. In addition of major mutation types which were detectable in direct sequencing analysis, all the patients showed to have minor types of mutations which were found only through BCR-ABL KD cloning and subsequent colony sequencing. To make sure that this minor mutation types were not caused by sequencing error, we also analyzed of 3 patients who showed TKI resistance, but had no BCR-ABL mutation. In addition, samples from 3 normal persons were analyzed with the same method. The frequency of appearance of the minor types of point mutation was reduced in the patient group who showed TKI resistance, but had no BCR-ABL mutation, and then dramatically decreased in the normal person group, indicating that BCR-ABL gene in patients with point mutation are relatively unstable. Analysis of serial samples from a same patient provided evidence of dynamic change of portion of compound mutation. In most case, portion of the clone containing compound mutation was increased as treatment went on, indicating the clone harboring compound mutation can take survival advantage over TKI treatment in comparison of the clone containing individual type of mutation. In addition, some patients showed change in individual mutation type comprising multiple mutation as treatment went on. Currently investigation of clinical relevance of compound mutation and other analyses are being carried on and more results will be provided in detail at the conference. Table 1. Patients Tx at mutation detection (mg) Compound type Compound % 1 Nilotinib400 G250E+T315I 6.7 G250E+D444G 33.3 T315I+D444G 6.7 2 Nilotinib400 M244V+T315I 95.0 3 Dasatinib100 Y253H+T315I 95.0 4 Dasatinib140 T315I+E459K 55.6 5 Dasatinib200 T315I+M351T 66.7 6 Dasatinib100 NCM Dasatinib80 NCM Dasatinib100 M244V+F359V 16.7 7 Bosutinib500 NCM 8 Dasatinib140 T315I+F359C 35.3 9 Imatinib400 E255K+T315I 5.6 10 Dasatinib80 E255V+T315I 90.0 11 Imatinib800 E255K+T315I 10.5 12 Nilotinib800 E255K+T315I 12.5 13 Dasatinib100 F311I+T315I 35.0 F311I+F317Lb 10.0 Imatinib400 F311I+T315I 10.0 F311I+F317La 15.0 F311I+F317Lb 55.0 14 Nilotinib800 Y253H+F359I 5.6 15 Bosutinib500 V299L+E459K 95.0 Nilotinib400 + Dasatinib100 V299L+F359I 5.0 V299L+E459K 55.0 V299L+F317La+E459K 15.0 V299L+F359I+E459K 15.0 V299L+F317La+F359I+E459K 5.0 16 Imatinib600 NCM 17 Imatinib400 NCM NCM: no compound mutation. Disclosures: No relevant conflicts of interest to declare.

BCR-ABL kinase domain mutation analysis in chronic myeloid leukemia patients treated with tyrosine kinase inhibitors: recommendations from an expert panel on behalf of European LeukemiaNet

Blood ◽  
2011 ◽  
Vol 118 (5) ◽  
pp. 1208-1215 ◽  
Author(s):  
Simona Soverini ◽  
Andreas Hochhaus ◽  
Franck E. Nicolini ◽  
Franz Gruber ◽  
Thoralf Lange ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Tyrosine Kinase ◽  
Tyrosine Kinase Inhibitors ◽  
Mutation Analysis ◽  
Myeloid Leukemia ◽  
Kinase Inhibitors ◽  
Direct Sequencing ◽  
Chronic Phase ◽  
Kinase Domain ◽  
Abl Kinase

AbstractMutations in the Bcr-Abl kinase domain may cause, or contribute to, resistance to tyrosine kinase inhibitors (TKIs) in chronic myeloid leukemia patients. Recommendations aimed to rationalize the use of BCR-ABL mutation testing in chronic myeloid leukemia have been compiled by a panel of experts appointed by the European LeukemiaNet (ELN) and European Treatment and Outcome Study and are here reported. Based on a critical review of the literature and, whenever necessary, on panelists' experience, key issues were identified and discussed concerning: (1) when to perform mutation analysis, (2) how to perform it, and (3) how to translate results into clinical practice. In chronic phase patients receiving imatinib first-line, mutation analysis is recommended only in case of failure or suboptimal response according to the ELN criteria. In imatinib-resistant patients receiving an alternative TKI, mutation analysis is recommended in case of hematologic or cytogenetic failure as provisionally defined by the ELN. The recommended methodology is direct sequencing, although it may be preceded by screening with other techniques, such as denaturing-high performance liquid chromatography. In all the cases outlined within this abstract, a positive result is an indication for therapeutic change. Some specific mutations weigh on TKI selection.

High Sensitive Detection of Imatinib-Resistance Associated Mutations Based on ARMS-PCR.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2943-2943
Author(s):  
Franz X.E. Gruber ◽  
Mikchail Soevershaev ◽  
Marita Olsen ◽  
Bjoern Skogen
Keyword(s):  
Kinase Inhibitors ◽  
Point Mutations ◽  
High Sensitivity ◽  
Kinase Domain ◽  
Single Step ◽  
Sensitive Detection ◽  
Imatinib Resistance ◽  
Abl Kinase ◽  
Loop Region ◽  
Highly Sensitive Detection

Abstract Background: Point mutations in the Abl kinase domain are associated with resistance against imatinib. Strategies to overcome resistance include dose escalation, combination treatment using imatinib with conventional or other developmental agents or, in the future, imatinib may be replaced by other tyrosine kinase inhibitors which work effectively against mutated clones. Mutational profiling of the BCR-ABL kinase domain will in this scenario become an important analysis as a supplement to BCR-ABL quantitation and may provide the rational basis for therapy, once resistance is diagnosed. Our group reported recently a sensitive, single step PCR assay for quantitation of mutated clones based on the ARMS principle. Aim: We describe an optimized, two step analysis for high sensitivity screening of mutated clones associated with resistance against imatinib targeting all P-Loop mutations, the T315I and M351T. Methods: In a first conventional PCR-reaction a cDNA-region spanning the BCR-ABL breakpoint is amplified resulting in an isolation of the BCR-ABL kinase domain for further analysis. An aliquot is then analysed in a second PCR step, conducted on the real time PCR Taqman platform. Selectivity for the mutated clone is conferred by the amplification refractoriness of non complementary primer 3′-ends (ARMS principle). By introducing potent nucleotide-mismatches in position n-2, selectivity of the assay could be further increased. Even in the P-Loop region, which is known to be a difficult PCR template, misannealing could be reduced to an acceptable level. Results: Assays targeting all P-Loop mutations inclusive the T315I and M351T were tested by analysis of patient samples diluted in normal cDNA and non-mutated BCR-ABL and plasmid dilutions, containing the targeted mutation in a background of wildtype plasmids. Generally a 1:1000 dilution of mutated templates could be detected (sensitivity 0.1%). For some mutations even higher sensitivity could be achieved (0.01%). The level of sensitivity is generally higher than reported for other methods described before. The first PCR step can be conducted in parallel to other PCR-based detection strategies. The second step can be run simultanously to Taqman based BCR-ABL quantitation. This makes the described assay the ideal supplement to general mutation detection approaches like D-HPLC or sequencing strategies. Compared to the single step assay we desribed before, the two step approach increases sensitivity with one or two log factors. Conclusion: The described assay may be suitable for highly sensitive detection of mutated clones in resistant CML patients as a supplement to less sensitive general screening approaches and BCR-ABL quantitation.

ShRNA Targeting p190BCR-ABL Successfully Eliminates Ph-ALL Cells with or without ABL Kinase Domain Mutation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1838-1838
Author(s):  
Muneyoshi Futami ◽  
Toshiyuki Hatano ◽  
Yasushi Soda ◽  
Seiichiro Kobayashi ◽  
Makoto Miyagishi ◽  
...  
Keyword(s):  
Lentiviral Vector ◽  
High Titer ◽  
Therapeutic Option ◽  
Lymphoblastic Leukemia ◽  
Philadelphia Chromosome ◽  
Kinase Domain ◽  
Leukemia Cells ◽  
Specific Cell ◽  
Dependent Manner ◽  
Abl Kinase

Abstract In the majority of Philadelphia chromosome (Ph)-positive acute lymphoblastic leukemia (Ph-ALL) cases, the resulting BCR-ABL gene generates 190 kD active tyrosine kinase (p190) which is responsible for leukemogenesis and can be a molecular target for therapy. Although a series of ABL kinase inhibitors including imatinib, nilotinib and dasatinib reveal potent activities against Ph-ALL, acquired resistance caused by point mutations in the kinase domain such as T315I still remains to be overcome. That is why a novel strategy is desired in the treatment of Ph-ALL. We previously reported that lentiviral delivery of maxizyme targeting p190 specifically induced apoptosis of Ph-ALL cells (Blood 104:356, 2004). Since RNA interference proved to be a more powerful tool in selective gene silencing, we applied this technology to test whether specific and efficient killing of Ph-ALL cells could be achieved by down-regulation of p190. We designed a series of 21-mer and 27-mer small hairpin RNA (shRNA) targeting p190 mRNA and constructed plasmid vectors expressing these shRNA, which were screened by transfection of 293T/p190 cells to determine optimal target sites. As a result, three candidate sequences were identified; junctional 27-mer, junctional 21-mer and ABL 21-mer. Then, we inserted each of the shRNA expression cassettes into the lentiviral vector (HIV-U6/shRNA) and prepared high titer virus stock for infection of leukemia cells. shBCR-ABL/21, but not shBCR-ABL/27, induced significant and specific cell death of p190+ Ph-ALL cells in a time-dependent manner. shABL was more potent than shBCR-ABL/21 and also active against p210+ CML cells as well as 293 cells, but did not substantially affect Ph-negative leukemia cells. Both shABL and shBCR-ABL/21 completely inhibited growth of Ba/F3 cells harboring either wild-type or mutant p190 which renders those resistant to imatinib. Furthermore, both shRNA at low multiplicity of infection additively cooperated with imatinib in growth inhibition of Ba/F3-p190 cells. These data suggest that shRNA targeting p190 may become a therapeutic option in Ph-ALL by improvement of its delivery system like liposome. Growth of BA/F3-p190BCR-ABL Cells transduced with shRNA Targeting p190BCR-ABL Growth of BA/F3-p190BCR-ABL Cells transduced with shRNA Targeting p190BCR-ABL

Sequencing of Subcloned PCR Products Facilitates Earlier Detection of BCR-ABL1T315I Mutants Compared to Direct Sequencing of the ABL1 Kinase Domain.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1952-1952
Author(s):  
Alfonso Quintás-Cardama ◽  
Don L. Gibbons ◽  
Hagop Kantarjian ◽  
Moshe Talpaz ◽  
Nicholas Donato ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Catalytic Domain ◽  
Direct Sequencing ◽  
Cytogenetic Response ◽  
Median Number ◽  
Kinase Domain ◽  
Therapeutic Implications ◽  
Abl Kinase ◽  
Therapeutic Algorithms ◽  
Pcr Products

Abstract ABL kinase domain mutations represent the most frequent cause of resistance to tyrosine kinase inhibitors (TKIs). The BCR-ABL1T315I mutation affects a highly conserved “gatekeeper” threonine near the ABL catalytic domain, thus causing steric hindrance that precludes ABL TKIs binding. BCR-ABL1T315I retains kinase activity even in the presence of micromolar concentrations of imatinib or dasatinib. Thus, early detection of BCR-ABL1T315I has important prognostic and therapeutic implications. We evaluated the sensitivity of detection of BCR-ABL1T315I in 62 CML pts after failure (n=51) or intolerance (n=11) of imatinib enrolled in a phase I study of dasatinib by direct sequencing (DS) of nested PCR-amplified BCR-ABL1 products as well as by DNA expansion of specific clones (DESC) followed by DNA sequencing of at least 10 clones. Ten (15%) pts were found to carry BCR-ABL1T315I, 4 prior to dasatinib start and 6 during dasatinib therapy. Four pts never responded to imatinib whereas 5 had achieved a complete hematologic response (CHR) and 1 a complete cytogenetic response (CCyR). Imatinib was stopped due to rash (n=1), hematologic resistance (n=4), and progression to accelerated (AP; n=1) or blastic (BP; n=4) phase. Of the 4 pts in whom BCR-ABL1T315I was detected by DESC prior to dasatinib start, 2 are dead and 2 are alive. DS performed on the same samples detected BCR-ABL1T315I only in 2 of them. In pts in whom direct sequencing failed to detect BCR-ABL1T315I, the percentage of clones carrying BCR-ABL1T315I was 10% and 100%, respectively. In one of them, DS detected BCR-ABL1T315I 12 months later, whereas in the other case, DS failed to detect BCR-ABL1T315I in 3 separate occasions. These 2 pts are still alive (1 on hydrea, 1 on bosutinib) but never achieved any cytogenetic response. BCR-ABL1T315I was detected in 6 additional pts after a median time of 5 months (range, 1–6) on dasatinib (dosing ranging from 70 to 140 mg/d): 5 of 5 analyzed by DESC and 1 of 2 determined by DS. Of them, 4 had no response to dasatinib and 2 had transient cytogenetic responses (1 minor, 1 partial) and only one is still alive. In 1 that had no response, for whom paired samples were available, BCR-ABL1T315I was present in 10% of clones but was not detected by DS. Overall, the median number of clones harboring BCR-ABL1T315I was 90% (range, 10%–100%) and the median number of mutants co-expressed with BCR-ABL1T315I was 3 (range, 1–9). DS failed to identify 40 non-BCR-ABL1T315I mutants (including F317L in 1 patient), regardless of the percentage of clones in which they were expressed, except for E355G in 1 patient. Eight pts received dasatinib for more than 3 weeks (median, 5 months; range, 2–13) and were evaluable for response. Six failed to achieve any cytogenetic response and 2 had transient cytogenetic responses (1 minor and 1 partial). Seven (70%) pts died and 3 are alive with no cytogenetic response. In conclusion, DS has a poor sensitivity to detect ABL kinase mutations, particularly when the proportion of mutated clones is low. With the advent of novel T315I inhibitors, prompt detection of this highly-resistant mutation must be prioritized and included in therapeutic algorithms. To maximize the sensitivity of T315I detection, sequencing of subcloned PCR products might be preferable to DS.

Treating Ph+ Acute Lymphoblastic Leukemia (ALL) in the Elderly: The Sequence of Two Tyrosine Kinase Inhibitors (TKI) (Nilotinib and Imatinib) Does Not Prevent Mutations and Relapse.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2601-2601 ◽  
Author(s):  
Cristina Papayannidis ◽  
Paola Fazi ◽  
Alfonso Piciocchi ◽  
Francesco Di Raimondo ◽  
Giovanni Pizzolo ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Tyrosine Kinase ◽  
Research Funding ◽  
Kinase Inhibitors ◽  
Lymphoblastic Leukemia ◽  
The Elderly ◽  
Kinase Domain ◽  
Intensive Chemotherapy ◽  
Molecular Responses ◽  
Abl Kinase

Abstract Abstract 2601 Background: Tyrosine Kinase Inhibitors (TKI) have been shown to be very effective for the treatment of Acute Lymphoblastic Leukemia (ALL), with a Complete Hematologic Remission (CHR) rate close to 100%, and a high rate of Complete Cytogenetic and Molecular responses (CCgR and CMR). However, when they are used alone, as single agents, most patients relapse, so that they are currently used in combination with chemotherapy and as a preparation to allogeneic stem cell transplantation (SCT). Since Ph+ ALL is more frequent in the elderly, many patients cannot tolerate intensive chemotherapy and are not eligible for SCT. We have explored if the administration of two TKIs, Nilotinib (NIL) and Imatinib (IM) can improve the results without increasing the toxicity. Aims: To evaluate the response and the outcome of Ph+ ALL patients treated with the sequential administration of NIL and IM, to investigate the type and number of BCR-ABL kinase domain mutations developing during and after the study. Methods: We have designed a study (ClinicalTrials.gov. NCT01025505) in which patients more than 60 years old or unfit for intensive chemotherapy and SCT where treated with two TKIs, NIL 400 mg twice daily, and IM 300 mg twice daily, alternating for 6 weeks for a minimum of 24 weeks (study core) and indefinitely in case of response. The 6-weeks rotation schedule was respected, irrespectively of temporary discontinuations. The primary end-point was the rate of Disease Free Survival (DFS) at 24 weeks (4 courses of treatment); the secondary end points included the evaluation of CHR, CCgR and CMR rates. Mutation analysis was performed by nested RT-PCR amplification of the ABL kinase domain of the BCR-ABL transcript (codons 206 through 421). Amplified products were screened by denaturing-high performance liquid chromatography (D-HPLC). Samples scored positive for the presence of sequence variations were then subjected to direct automatic sequencing to characterize the mutation. Results: 39 patients have been enrolled in 15 Italian hematologic Centers (median age 66 years, range 28–84). Among these, 8 patients were unfit for standard chemotherapy or SCT (median age 50 years, range 28–59). 27 patients were p190, 5 were p210 and 7 were p190/p210. After 6 weeks of treatment, 36 patients were evaluable for response: 34 were in CHR (94%) and 2 in PHR (6%). 23 patients have already completed the study core (24 weeks), 87% were in CHR and 17 are currently continuing therapy in the protocol extension phase. Thus, the OS at 1 year is 79%, and 64% at 2 years. Overall, 1 patient was primarily resistant and 13 patients have relapsed, with a median time to relapse of 7.6 months (range 0.8–16.1 months), for a DFS of 51.3% at 12 months (Figure 1). Mutations detected were T315I in 2 cases, Y253H in 3 cases, T315I and Y253H in 1 case, E255K in 1 case, T315I and E255K in 1 case, E255V and Y253H in 1 case. Two patients were WT. A detailed kinetics of Molecular responses is shown in Table 1. Data on mutational analysis are reported in Table 2. Further details about Cytogenetic and Molecular responses, and about Adverse Events will be provided on site. Conclusions: In this small cohort of Ph+ ALL elderly/unfit patients, the rates of relapse and progression were not likely to be different from the rates observed with Imatinib alone (Vignetti et al, Blood 2007, May 1;109(9):3676-8) and Dasatinib alone (Foà, Blood 2011, Dec 15;118(25):6521-8). It's important to notice that the mutations that occurred at the time of relapse were sensitive to other TKIs (Dasatinib and Ponatinib). Acknowledgments: COFIN, Bologna University, BolognAIL, PRIN, Fondazione del Monte di Bologna e Ravenna, INPDAP. Disclosures: Pizzolo: Hoffmann-La Roche: Consultancy, Honoraria. Luppi:CELGENE CORPORATION: Research Funding. Vallisa:CELGENE CORPORATION: Research Funding. Martinelli:NOVARTIS: Consultancy, Honoraria, Speakers Bureau; BMS: Consultancy, Honoraria, Speakers Bureau; PFIZER: Consultancy; ARIAD: Consultancy. Baccarani:ARIAD, Novartis, Bristol Myers-Squibb, and Pfizer: Consultancy, Honoraria, Speakers Bureau.

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